DESCRIPTION: (Applicant's Description) DNA double-strand breaks (DSBs) occur spontaneously and are caused by exposures to a variety of environmental insults including ionizing radiation, oxidative stress, metals, and natural and man-made genotoxic substances. Accurate repair of DSBs is essential for cell survival and for the prevention of genome rearrangements that may lead to cancer. Human somatic cells have several mechanisms for repairing DSBs, but the most commonly used mechanism appears to be the error prone non-homologous end-joining pathway (NHEJ). This pathway includes at least five proteins that appear to be unique to the NHEJ mechanism, the three components of DNA-PK (DNA-PKcs, Ku70, Ku80), the double-stranded DNA activated protein kinase, and DNA ligase IV and XRCC4. Additional NHEJ components (e.g. MRE11, polymerase b, exonuclease FEN-1) may be shared with other repair systems, and undoubtedly other NHEJ components remain to be discovered. Together the genes for the five unique NHEJ components, which must work in concert, span about one megabase of the human genome, 20,145 bp of which code for amino acids. Natural genetic variation is expected to yield about20 single nucleotide variants within coding regions of this gene set, of which about half might be expected to alter amino acid sequences. Half of these will occur within the very large DNA-PK catalytic subunit, DNA-PKcs. We propose to use denaturing high-performance liquid chromatography and direct nucleotide sequencing of PCR products to discover the common polymorphisms in all ( about160) of the exons of the five NHEJ genes from about 90 different reference individuals. Then we will examine the pattern of polymorphisms in patients with medical conditions that might suggest a deficiency in the activity of one or more of these genes. Finally, we will characterize individual reactions within the NHEJ pathway in cell lines from individuals with different NHEJ polymorphims to determine if the efficiency or accuracy of NHEJ in humans is affected by the natural genetic variations within these genes in a way that might contribute to human disease. This study also will provide a moderately dense set of mapped polymorphic markers within the five know NHEJ genes for subsequence linkage studies that could reveal subtle effects and more complex allele interactions.